ABSTRACT This application addresses PAR-18-042, Ancillary Studies to Major Ongoing Clinical Research Studies to Advance Areas of Scientific Interest within the Mission of the NIDDK, through a prospective study of diet, lifestyle and the gut microbiome in a large, prospective cohort. Diverticulitis is one of the most common gastrointestinal indications for hospitalization in the U.S. as well as a frequent indication for outpatient clinic visits. At least 20% of individuals with an initial episode will have one or more recurrences. Currently, there is no proven medical means to prevent diverticulitis. Several lines of evidence support the scientific premise that the interaction between diet and lifestyle factors and the gut microbiome plays a critical role in the etiopathogenesis of diverticulitis. However, no study has prospectively examined the relationship between diet, lifestyle, the gut microbiome and diverticulitis risk, or used metagenomic or metabolomic analysis to elucidate microbial genes, pathways and products important in this disease. Prospective studies are critical to offer rigorous data to infer causality. Symptoms are likely to subsequently influence diet and lifestyle, which, along with disease-associated inflammation and antibiotics, can change gut microbial profiles. Cross-sectional studies are thus unable to distinguish if lifestyle or microbial associations are a consequence or cause of the disease. Thus, we propose a first-of-its-kind large, prospective study to examine diet, lifestyle, and the gut microbiome/metabolome, in relation to incident and recurrent diverticulitis. In this ancillary study, we will leverage ongoing, already supported collection of stool samples from 35,000 participants enrolled in the Nurses? Health Study (NHS) II. This cohort of female nurses aged 25-42 years were enrolled in 1989 and have been followed biennially with validated assessments of diet, lifestyle, and medical history with >90% follow-up. After stool collection, we will follow the cohort for incident and recurrent diverticulitis and collect new data through a follow-up stool sample and post- diagnostic diet/lifestyle survey. All cases of diverticulitis will be confirmed via record review. To characterize the gut microbiome and metabolome, we will utilize cutting-edge platforms and analytic pathways. Prospectively collected, pre-diagnostic stool samples from a large cohort of individuals with validated data on clinical, dietary and lifestyle exposure over extended follow-up provide an unprecedented, cost-effective opportunity to examine the relationship between the gut microbiome, metabolome, diet and lifestyle factors, and diverticulitis. This study is critical to understanding the pathogenesis of this poorly understood condition with high economic and clinical burden, laying the groundwork for clinical trials of predictive biomarkers and lifestyle modification or therapeutic manipulation of gut microbes for prevention of complications and recurrence. Finally, as diverticulitis represents a clear manifestation of a perturbation in host-microbial interaction as a mechanism of disease, our study has implications for other microbiome and mucosal immune-mediated conditions.